Abstract Equine-assisted services (EAS) are known to be beneficial for participants, though factors that influence their in-session experiences of horse-human interactions (HHI) remain underexplored. A mixed-methods, repeated measures design was used to investigate relationships between unmounted activities and human mood state, their activity enjoyment, and their perceptions of the horse they interacted with. We aimed to determine if participants’ ( n = 56) reported mood states reflected their behavior toward the EAS horses ( n = 14) across activity conditions. Horses and humans completed three activity conditions: grooming, leading the horse, and leading the horse around a maze. To assess participants’ mood states throughout the interaction, we measured participants’ mood states before the interaction began and after each activity. A post-interaction survey was used to assess participants’ enjoyment of each activity and the influence of certain horse characteristics on their experience. All interaction activities were filmed for analysis of human behavior. Results from two logistic regression models suggest that activity type was not significantly associated with the arousal ( p = 0.09) or valence ( p = 0.70) of participants’ reported mood states; however, a trend in the post hoc comparisons indicated participants were more likely to report an activated mood state before the interaction began compared to after the first activity (i.e., post-groom) (OR: 3.15, CI: 7.79−1.28, p = 0.013). Most participants ( n = 46/54) reported pleasant moods throughout the interactions, although qualitative results suggest participants experienced a wider range of affective states. Participant behavior supported their reported pleasant mood as most behaviors across conditions were affiliative toward the horse. These results identify factors that influence human experiences when interacting with horses that are incorporated into EAS.